2. CELL CYCLE AND APOPTOSIS
Cells undergo 3 controlled processes
The first two are part of the cell cycle, the last an exit from the cell cycle
Division (the cell cycle)
Quiescence
This is where most of the work of being a cell lies
During division the energy of the cell is devoted to making a new cell
Death
This can be a normal process creating a final functional form or an induced suicide
Epithelium and reticuloendothelial cells undergo active transitions towards terminally
differentiated states in which the final forms are unable to divide
The stratum corneum consists of cells that have become bags of crosslinked keratin protein with
no internal metabolism
Suicide can be induced because the organism senses a threat to the entire organism
Infection, cancer, avoidance of autoimmunity
17BCB0087 A.G.Harith Laxman
3. CONTROL OF ENTRY INTO CELL CYCLE
AND APOPTOSIS
Cell cycle is initiated by
phosphorylation of transcription
factors
These activate transcription of a set of
proteins known as cyclins
The appearance of cyclins is
progressive and determines the types
of proteins that will be phosphorylated
at a particular point during the cell
cycle
17BCB0087 A.G.Harith Laxman
4. CYCLINS AND CDK’S CDK levels don’t change while
cyclins are destroyed at the end
of each phase
There are 3 general groups of
each
G1 cyclins
Cyclin D
S-phase cyclins
Cyclin A
G2 cyclins
Cyclin B (maturation promoting
factor MPF)
Cyclin E is shared between G1
and M phase
Cyclin A is shared between M
phase and G2
17BCB0087 A.G.Harith Laxman
5. CYCLINS BIND
CDK’S
CDK’s are Cyclin Dependent Kinases
Association with cyclins activates
their kinase function
A cyclin tethers a target protein to the
CDK
The targets of CDK’s are
transcription factors among other
proteins
CDK’s are serine/threonine kinases
17BCB0087 A.G.Harith Laxman
6. THE EXIT FROM G0
G0 is a quiescent state
Activation of G1 CDK occurs due to a
rising level of G1 cyclins
G1 cyclins are transcriptionally activated
by growth factors
17BCB0087 A.G.Harith Laxman
7. EVENTS DURING G1
A rising level of G1 cyclins increases
the activity of G1 CDK’s
CDK’s in turn activate proteins and in
turn genes that prepare the cell to
begin DNA replication
At the G1 S boundary, the cell
encounters a checkpoint
17BCB0087 A.G.Harith Laxman
8. G1/S
CHECKPOINT
This is controlled by the activity of the
transcription factor E2F
E2F is a family of related proteins (E2F 1 to E2F5)
E2F is found complexed throughout the cell cycle
to another family of proteins: Rb
At the G1/S checkpoint, Rb is phosphorylated by
CDK2/cyclinA
E2F is freed from sequestration and activates
transcription at genes containing an E2F consensus
sequence
17BCB0087 A.G.Harith Laxman
9. AND THOSE GENES ARE
Three groups
Cell cycle regulators
Cyclin A
E2F, Rb, myc, myb
Note that these are not all positive
regulators of cell cycle
Enzymatic machinery for DNA
synthesis
DNA polymerase
PCNA
Enzymes involved in nucleotide
metabolism
DNA synthesis regulators
Enzymes that recognize the origins
of replication for example17BCB0087 A.G.Harith Laxman
10. OTHER CHECKPOINTS
These monitor the completion of
DNA synthesis
The presence of Okazaki fragments
prevents entry into G2
DNA damage
This occurs before, during and after
completion of S phase
Spindle attachment
Failure to attach spindle to
centromere results in blockage of
mitosis at metaphase
Failure to align the spindle during
cytokinesis results in blockage at
anaphase17BCB0087 A.G.Harith Laxman
11. DOWNREGULATION OF CYCLIN INFLUENCED CDK ACTIVITY
cyclin B
cyclin A
ribonucleotide
reductase
Mitosis MitosisInterphase Interphase
Time
Newly synthesized proteins labeled with 35S-methionine:
This is accomplished through proteolysis
of the cyclins
G1 phase cyclins disappear during S and G2
phase
M-phase promoting factor (CDK2 + cyclin
B) concentrations rise just prior to onset of
mitosis
Cyclins associated with MPF are degraded by
anaphase promoting complex
Cyclin B levels peak at G1/M
Degradation during anaphase
APC promotes polyubiquitination of cyclin B
Ubiquitinated cyclin B is degraded by a
proteosome
Cyclin transcription is also turned off and
the mRNA is unstable
So no new cyclin is made until transcription
is restored
17BCB0087 A.G.Harith Laxman
13. IN THE OVERALL
Stimulated entry into G1 results in
appearance of an initial level of
cyclins that promote the progressive
activation of genes enabling the cell
to synthesize DNA
A series of progressive steps result in
Activation of genes further into the
cycle
Degradation of cyclins that promoted
earlier steps
Passage through checkpoints that
insure mechanistic fidelity of each step
17BCB0087 A.G.Harith Laxman
14. APOPTOSIS
This is programmed cell death
Distinguish it from necrosis
Necrosis results from traumatic forces
outside the cell
Necrotic tissue provokes
inflammation as the immune system
moves in to clear out damaged and
dead cells
Apoptosis is an ordered stepwise
self-destruction that permits
surrounding cells to utilize the
breakdown products of the dead
cell
There is no inflammation involved17BCB0087 A.G.Harith Laxman
15. THE APOPTOTIC CELL
Mitochondria break open
DNA fragments in a regular way
The cell loses a regular shape
Undergoes blebbing
This is an irregular bubbling appearance of
the plasma membrane
17BCB0087 A.G.Harith Laxman
16. THE MECHANISMS OF APOTOSIS
Can be classified as externally or
internally signaled
One internal route involves p53
p53 is a transcription factor that is
involved in cell cycle control and
sensing the presence of DNA damage
The central role p53 plays is at the
G1/S checkpoint
17BCB0087 A.G.Harith Laxman
17. P53 CONTROLS ENTRY INTO S-
PHASE
P53 can sense DNA damage by binding
mismatches
In the presence of damage, p53 activates
transcription of p21
P21 binds and inactivates CDK2-cyclin E complexes
The complex is unable to phosphorylate Rb and free E2F
Thus entry into S phase is inhibited
If the damage is repaired, p53 levels and p21 levels drop
and S phase ensues
17BCB0087 A.G.Harith Laxman
18. BUT IF THE DNA DAMAGE IS
EXTENSIVE
P53 induces apotosis by activating transcription of Bax
BAX protein competes with BCL-2 to form pores in
mitochondrial membranes
BCL-2 prevents the release of cytochrome c from mitochondria into the
cytoplasm
BAX permits release of cytochrome c
When released, cytochrome c stimulates caspase activation
17BCB0087 A.G.Harith Laxman
19. THE CASPASES
These are proteolytic enzymes that
are held in check by external or
internal inhibitors
Activation results in an explosive
proteolytic cascade
Caspase 9 cleaves and activates
other caspases
The caspases also activate
quiescent nucleases
17BCB0087 A.G.Harith Laxman
20. EXTERNAL APOPTOTIC MECHANISMS
Involve external “death signals”
Cells may be recognized as a threat to
the whole organism
The immune system moves in to kill them
One mechanism of killing involves a
command to the cell to initiate apoptosis
17BCB0087 A.G.Harith Laxman
21. FAS/FAS LIGAND SIGNALING
Fas ligand (FasL) is a membrane
bound cell surface protein
It binds to Fas receptor
Binding results in trimerization
and activation of APAF
APAF in turn activates caspase 8
by proteolysis of a caspase 8
zymogen
Caspase 8 cleaves a BCL-2 family
member BID
BID translocates to the mitochondria
and binds BAX
Bax permits leakage of cytochrome
c and activation of the caspase 9
cascade via APAF-1 again
17BCB0087 A.G.Harith Laxman
